This research proposal is an experimenal investigation into (1) the identification of synaptic transmitters, (2) the physiological basis of transmitter action at the single cell level, and (3) the morphological identification of physiologically and pharmacologically characterized cells. Inhibitory and excitatory transmitter agonists and antagonists are added to the bathing medium using perfused-retina eyecup preparations of rabbits and several types of amphibians (mudpuppy and tiger salamander). Analysis of drug action is based on intracellular electrode experiments using conventional recording techniques, single electrode voltage clamp techniques ad a lock-in amplifier for quantitative measurements of conductance changes. A similar, parallel set of experiments will be carried out using enzyme dissociated cells maintained in tissue culture conditions. Intracellular recording of single dissociated cells will be analyzed as agonists/antagonists are added to the bathing medium, iontophoretically applied, or introduced through pressure injection. The isolated retina of amphibians and rabbits will be studied for types and levels of amino acids released into the bathing medium, using reverse-phase high performance liquid chromatography. Particular emphasis will be placed on the identity of rod and cone transmitters. These experiments will also be combined with tissue culture techniques to fractionate the retina into limited neuronal populations, through the use of neuroactive ligands. The goal of this research is to identify synaptic transmitters and associate them with specific pathways and cell types. Several disease states which afflict the retina (retinitis pigmentosa; macular degeneration), show prolonged states in which photoreceptor function is compromised while leaving a more or less intact retina. An understanding of the chemicals involved in photoreceptor transmission, together with the synaptic receptors which mediate their action may help in the design of drug therapy to assist patients with limited vision.
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